Suspicion Of Heparin-induced Thrombocytopenia Research Articles

Is the combination of two automated rapid assays for diagnosis of heparin-induced thrombocytopenia necessary?

Heparin-induced thrombocytopenia (HIT) is an immune-mediated condition characterized by a decrease in platelet count and an increased thrombotic risk. HIT event is caused by antiplatelet factor/heparin (PF4/H) antibodies that can activate the platelets. The diagnosis of HIT is based on a clinical evaluation and laboratory results. Aim of our study was to evaluate whether the combined use of two rapid assays for diagnosis of HIT provides a diagnostic advantage over the use of a single automate assay. We extracted from the laboratory informatic system all the determinations requested for the detection of antibodies against heparin/PF4 complexes from July 2020 to June 2024 (n. 229). In our laboratory, total antibodies against heparin/PF4 complex [HemosIL HIT-Ab-(PF4-H), Instrumentation Laboratory] and IgG Ab (HemosIL AcuStar HIT-IgG, Instrumentation Laboratory) are measured simultaneously with two different instruments. Two hundred six samples tested negative for both methods, 23 samples tested positive for at least one method and nine samples tested positive for both methods. The grade of concordance between the two assays shows a weighted Kappa of 0.536 (moderate agreement). No sample tested positive only for IgG-Ab. The sensitivity of HIT-Ab-(PF4-H) was 1, whereas the specificity was 0.95. For the HIT-IgG (PF4-H) method, sensibility and specificity were 0.77 and 1, respectively. Our results suggest that performing these two tests simultaneously does not provide additional useful information in patients with suspicion of HIT. The total Ab assay seems to be sufficient, as it shows higher sensitivity although at the expense of lower specificity.

Hematological features and alternate diagnoses in critically ill thrombotic antiphospholipid syndrome patients.

Severe thrombotic antiphospholipid syndrome (APS) frequently affects the kidney, heart, and central nervous system. The precise frequency, clinical picture, differential diagnoses, and outcome of APS-related hematological involvement are lacking, especially in patients requiring ICU admission. This study aimed to describe the hematological manifestations associated with critically ill thrombotic APS patients and catastrophic antiphospholipid syndrome. This French, national, multicenter, retrospective study, conducted, from January 2000 to September 2018, included all APS patients admitted to 24 participating centers' ICUs with any new thrombotic manifestation. The prevalence of hematological manifestations and their associated outcomes were studied. One hundred and thirty-four patients, female 72%, median [IQR] age 45 [34-56] years, with 152 episodes were included. Anemia was present in 95% of episodes and thrombocytopenia in 93%. The lowest values for hemoglobin and platelets were 7.1 [6.3-8.8] g/dL and 38 [21-60] g/L, respectively. The lowest platelet count below 20g/L was significantly associated with a higher in-ICU mortality rate (50%, p < 0.0001). A thrombotic microangiopathy syndrome (TMA) syndrome was seen in 16 patients (12%) and was associated with higher in-hospital mortality (p = 0.05). Median ADAMTS-13 levels were 44% [27-74]. Anti-ADAMTS13 antibodies were tested in 11 patients and found negative in all. A suspicion of heparin-induced thrombocytopenia (HIT) was raised in 66 patients but only four patients were classified as definite HIT. Disseminated intravascular coagulation (DIC) was seen in 51% of patients. Thrombocytopenia is very frequent in severe APS patients and may be related to TMA, HIT, or DIC. Deciphering the mechanisms of thrombocytopenia is decisive in CAPS patients. Key Points • Thrombocytopenia is the hallmark laboratory finding in CAPS. • A complete thrombotic microangiopathy pattern is infrequent in CAPS patients. • Alternate diagnoses of CAPS, especially heparin-induced thrombocytopenia, need to be adequately investigated.

Association of Metal Cations with the Anti-PF4/Heparin Antibody Response in Heparin-Induced Thrombocytopenia.

Heparin-induced thrombocytopenia (HIT) is an antibody-mediated immune response against complexes of heparin and platelet factor 4 (PF4). The electrostatic interaction between heparin and PF4 is critical for the anti-PF4/heparin antibody response seen in HIT. The binding of metal cations to heparin induces conformational changes and charge neutralization of the heparin molecule, and cation-heparin binding can modulate the specificity and affinity for heparin-binding partners. However, the effects of metal cation binding to heparin in the context of anti-PF4/heparin antibody response have not been determined. Here, we utilized inductively coupled plasma mass spectrometry (ICP-MS) to quantify 16 metal cations in patient plasma and tested for correlation with anti-PF4/heparin IgG levels and platelet count after clinical suspicion of HIT in a cohort of heparin-treated patients. The average age of the cohort (n = 32) was 60.53 (SD = 14.31) years old, had a mean anti-PF4/heparin antibody optical density [OD405] of 0.93 (SD = 1.21) units, and was primarily female (n = 23). Patients with positive anti-PF4/heparin antibody test results (OD405 ≥ 0.5 units) were younger, had increased weight and BMI, and were more likely to have a positive serotonin release assay (SRA) result compared to antibody-negative patients. We observed statistical differences between antibody-positive and -negative groups for sodium and aluminum and significant correlations of anti-PF4/heparin antibody levels with sodium and silver. While differences in sodium concentrations were associated with antibody-positive status and correlated with antibody levels, no replication was performed. Additional studies are warranted to confirm our observed association, including in vitro binding studies and larger observational cohorts.

Combination of 2 Quantitative Immunoassays and Clinical Score Algorithm to Reduce False-Negative Results in Heparin-Induced Thrombocytopenia: Prevalence Study of Mauriziano Hospital in Turin, Italy.

Heparin-induced thrombocytopenia (HIT) is a serious adverse effect of heparin treatment caused by platelet-activating anti-platelet factor 4 (PF4)/heparin antibodies. Accurate diagnosis of HIT is essential but remains challenging. The aim of our study was to explore the performance of our optimized diagnostic laboratory algorithm, based on Chemiluminescence (CliA) and ELISA immunoassays, on suspected HIT patients. The study compared the prevalence of HIT diagnosis in A.O. Mauriziano with the literature. 163 consecutive patients were investigated for suspected HIT with CliA HemosIL Acustar HIT-IgG, Werfen. HIT was ruled out in all patients with CliA <0.13 U/mL. All patients with CliA >0.13 U/mL were further investigated with Zymutest-HIA anti-PF4 IgG ELISA immunoassay. In these patients, HIT was ruled out on the combination of CliA between 0.13 and 1.0 U/mL followed by ELISA assay <0.300 OD. HIT was ruled in patients whose plasma tested positive or doubtful with CliA and positive with ELISA immunoassay and confirmed positive with a platelet aggregation test (PAT). Suspicion of HIT was revealed with clinical 4Ts score or recent suggestive anamnestic history. Our diagnostic algorithm ruled out HIT diagnosis in 144/163 patients (88%) and predicted a positive PAT in 5/19 (26%) of CliA positive (4/5) or ELISA positive and CliA doubtful (1/5) patients. Our prevalence was 3.1%, comparable with the literature. The approach combining 2 quantitative immunoassays' (CliA and ELISA) results and 4Ts score probability was able to rule out the diagnosis within 1 h in 66% of patients with suspected HIT and within 24 h in 88% of patients. In the remaining 12% of cases, management decisions have to be based on individualized judgment while awaiting functional confirming results (48-72 h).

Association of Metal Cations with the Anti-PF4/Heparin Antibody Response in Heparin-Induced Thrombocytopenia.

Heparin-induced thrombocytopenia (HIT) is an antibody-mediated immune response against complexes of heparin and platelet factor 4 (PF4). The electrostatic interaction between heparin and PF4 is critical for the anti-PF4/heparin antibody response seen in HIT. The binding of metal cations to heparin induces conformational changes and charge neutralization of the heparin molecule, and cation-heparin binding can modulate the specificity and affinity for heparin-binding partners. However, the effects of metal cation binding to heparin in the context of anti-PF4/heparin antibody response have not been determined. Here, we utilized inductively coupled plasma mass spectrometry (ICP-MS) to quantify 16 metal cations in patient plasma and tested for correlation with anti-PF4/heparin IgG levels and platelet count after clinical suspicion of HIT in a cohort of heparin-treated patients. The average age of the cohort (n = 32) was 60.53 (SD = 14.31) years old, had a mean anti-PF4/heparin antibody optical density [OD405] of 0.93 (SD = 1.21) units and was primarily female (n = 23). Patients with positive anti-PF4/heparin antibody test results (OD405 ≥ 0.5 units) were younger, had increased weight and BMI, and were more likely to have a positive serotonin release assay (SRA) result compared to antibody negative patients. We observed statistical differences between antibody positive and negative groups for sodium and aluminum and significant correlations of anti-PF4/heparin antibody levels with sodium and silver. While differences in sodium concentrations were associated with antibody positive status and correlated with antibody levels, no replication was performed. Additional studies are warranted to confirm our observed association, including in vitro binding studies and larger observational cohorts.

Accuracy of Diagnosing Heparin-Induced Thrombocytopenia

Heparin-induced thrombocytopenia (HIT) is a life-threatening condition that requires urgent diagnostic clarification. However, knowledge of the diagnostic utility of the recommended diagnostic tests is limited in clinical practice. To evaluate the current diagnostic practice for managing the suspicion of HIT. This prospective diagnostic study was conducted from January 2018 to May 2021 among consecutive patients with suspected HIT from 11 study centers in Switzerland, Germany, and the United States. Detailed clinical data and laboratory information were recorded. Platelet factor 4/heparin antibodies were quantified using an automated chemiluminescent immunoassay (CLIA). A washed-platelet heparin-induced platelet activation (HIPA) test was used as a reference standard to define HIT. Suspicion of HIT. The primary outcome was the diagnostic accuracy of the 4Ts score, the CLIA, and the recommended algorithm serially combining both tests. Of 1448 patients included between 2018 and 2021, 1318 were available for the current analysis (median [IQR] age, 67 [57-75] years; 849 [64.6%] male). HIPA was positive in 111 patients (prevalence, 8.4%). The most frequent setting was intensive care unit (487 [37.0%]) or cardiovascular surgery (434 [33.0%]). The 4Ts score was low risk in 625 patients (46.8%). By 2 × 2 table, the numbers of patients with false-negative results were 10 (9.0%; 4Ts score), 5 (4.5%; CLIA), and 15 (13.5%; recommended diagnostic algorithm). The numbers of patients with false-positive results were 592 (49.0%; 4Ts score), 73 (6.0%; CLIA), and 50 (4.1%; recommended diagnostic algorithm), respectively. In this diagnostic study of patients suspected of having HIT, when the recommended diagnostic algorithm was used in clinical practice, antibody testing was required in half the patients. A substantial number of patients were, however, still misclassified, which could lead to delayed diagnosis or overtreatment. Development of improved diagnostic algorithms for HIT diagnosis should be pursued.

Implementation of 4T Score Documentation on PF4 Testing in Patients with Suspected Heparin-Induced Thrombocytopenia

Purpose: Heparin-induced thrombocytopenia (HIT) is a severe complication of heparin therapy. Diagnosis requires clinical evaluation and laboratory testing. The 4T score is a pretest clinical scoring tool used to determine the probability of HIT. A non-profit health system implemented mandatory documentation of 4T score calculation prior to ordering PF4 tests for suspected HIT patients. The purpose of this performance improvement project is to evaluate the implementation of 4T score documentation on PF4 testing and argatroban utilization. Methods: This was a retrospective, multi-site chart review of patients conducted to assess the impact of mandatory 4T score documentation prior to a PF4 order. The study included adult patients who were admitted to one of the practice sites between January 2020 and November 2022 with a suspicion of HIT and a PF4 order. The primary outcomes included the number of PF4 tests ordered and the percentage of positive PF4 results. Secondary outcomes included number of SRA orders and percentage of positive SRA results, turnaround time for SRA, and the utilization of argatroban. Data collection included baseline demographics, documentation of 4T score, PF4 results, SRA results, documentation of heparin allergy, initial anticoagulant prescribed, platelet trend, ordering provider, and duration of argatroban therapy in patients with a positive PF4 result. The analysis involved using χ2 test and Fisher's exact test for the primary outcomes. Results: Mandatory 4T score documentation significantly reduced PF4 orders by 68% over a two-year period (p&amp;lt;0.0001). It also improved the percentage of positive PF4 results from 12% to 24% (p=0.60) and positive SRA results from 19% to 44% (p=0.35). Notably, after documentation of the mandatory 4T score tool, only 21% of the patients had a PF4 order placed. A total of 42 patients with a positive PF4 test were included to assess argatroban utilization. The turnaround time for SRA in these patients was 3.6 days while duration of argatroban therapy was 6.8 days. Conclusion: Implementation of mandatory 4T score documentation significantly reduced PF4 orders and improved HIT diagnostic testing in a non-profit health system. Challenges remain with SRA turnaround time and corresponding duration of argatroban therapy. The results of this project have demonstrated that standardized documentation is essential for optimizing HIT diagnosis and management protocols. Future efforts will focus on optimizing confirmatory HIT testing and non-heparin anticoagulation therapy.

The Impact of Laboratory Sample Request Evaluation and the Use of Probability Testing for HIT By Trained Physician on the Decrease of Unnecessary Tests and Increase the Likelihood of Positivity

Introduction: Heparin-induced thrombocytopenia (HIT) is an extremely serious and potentially fatal condition that can develop in patients taking heparin-based medications, such as unfractionated heparin (UFH) or low-molecular-weight heparin (LMWH). The laboratory tests for diagnosis of HIT is expensive and time consuming. Clinical assessment (4Ts) followed by testing for Heparin/platelet factor 4 antibody in intermediate and high risk patients is the standard algorithm of pretest for (HIT) which can be later confirmed by serotonin releasing assay if available. Materials and methods: We conducted a retrospective analysis of laboratory practice of pretest evaluation of samples submitted for HIT testing in a cohort of patients treated in a tertiary hospital in Saudi Arabia who had a clinical suspicion of HIT. The 4 T score was a pretest requisite to proceed for electronic request for HIT. It had to be performed by the requesting physicians of different medical specialties. The request then been evaluated by hematopathologist before proceeding. The test will be cancelled if: the 4T score calculated by attending physician is &amp;lt; 4 or if platelet count is normal with no evidence of significant drop in the last 4 days from request or the test was performed at least 2 days before the new request. Additional exercise was performed by two physicians in training after train them to standardize the calculation of the 4T scores without re evaluation by the hematopatologist. The sample proceeded for testing if it met the testing criteria of 4T score &amp;gt; 3 and passed the evaluation by the hematopathologist. All tested sample were performed by immunoglobulin G (IgG) antibodies via an ELISA (Asserachrom HPIA-IgG, Stago Diagnostica STA®) and Optic density ( OD) and was evaluated and considered positive if OD &amp;gt;0.39 with sub classification to low positive 0.4-1, moderate 1-2 and strong if &amp;gt;2. Over a period of 2 years, we received 449 samples for patients who were admitted to our hospital, received prophylactic or therapeutic doses of LMWH or UFH, and underwent an HIT test due to clinical suspicion ( &amp;gt; 50% drop in platelet count from baseline, or ≥ 2 consecutive platelet counts &amp;lt; 150,000 per mm 3 and was defined as possible HIT). The 4 T score was calculated in the hospital system by attending physicians referred to as (system 4T score) and recalculated by trained two physicians and referred to (re calculated 4T score). Descriptive statistics, paired samples statistics, correlations, and analysis of variance (ANOVA) were performed. Results: The test was labeled by the laboratory physician ( hematopathologist ) as not indicated and test was not performed in 156 (35%) for various reasons including low 4 T score &amp;lt;3 in 90 (57%), Normal platelet count &amp;gt; 150 at time of test ( and wrong calculated 4T score) in 28 (18%), duplication of test order in 10 (6.4%%), There was clear other cause of thrombocytopenia in 15 (10%), Bad sample quality 5 (3.2%%) and 8 (5%) for other reasons. The samples for HIT test were for 231(51%) males and 220 (49%) females with mean age 56.3 (1-102) years. The mean platelet count at time of request was 82.9 (2-384) per mm 3. Out of the total cohort 422 (95.1%) patients had received UFH and only 22 (4.9%) received LMWH. There is only 69/449 (15.4%) agreement between the two calculations (system and re calculated score) . Out of the tested samples 293, total positive tests 19 (6.5%), were 11 (3.8%) had low positive for HIT test, moderate positive in 6 (2%) and only 2 strongly positive for HIT. The calculated (system 4T score) of &amp;lt; 4 were 142 (32%) samples while the (re calculated 4T score) were 136 (30%) of which 60 ( 38%) of the cancelled tests by hematopathologist, of which 68 (23%) of tested samples were negative and 7 (37%) of positive tests could have been missed based on this calculation. There were 12 out of 19 had moderate and high score by calculation and positive HIT while there were 17 out of 19 had moderate and high score by system calculation. Conclusion: We are reporting improved detection of HIT and confirmed the value of applying the 4 T score for HIT testing with added value of pre test screening to exclude low probability and cancel samples with no clear indication of HIT due to presence of other causes of thrombocytopenia.The retrospective re calculation even with more trained physicians did not add a value compared to pretest evaluation and screening by hematopathologist which had resulted in cancelation or more than third of the requested tests.

Cerebral venous sinus thrombosis associated with SRA-negative heparin-induced thrombocytopenia: case report

BackgroundThere are very few documented reports in literature of cerebral venous sinus thrombosis (CVST) caused by immune-mediated heparin-induced thrombocytopenia (HIT). Further, there are very few reports of false negative serotonin release assays (SRAs) when testing for immune-mediated HIT.Case presentationWe present a case of a 60- year-old male with recent unfractionated heparin administration for venous thromboembolism prophylaxis, an elevated 4T score of 5 and acute CVST in which immune-mediated HIT was suspected. The enzyme-linked immunosorbent assay (ELISA) screening assay was positive for PF4 antibodies and subsequent reflexive SRA testing was negative. However, given the clinical picture, a false-negative SRA was suspected (and eventually confirmed), prompting use of the alternative PF4-dependent p-selectin expression assay (PEA) which was confirmed to be positive. The patient was successfully managed with a bivalirudin infusion and eventually transitioned to apixaban.ConclusionIt is uncommon for immune-mediated HIT with thrombosis to manifest as CVST. Similarly, false-negative SRA is uncommon in immune-mediated HIT. Take-away lessons from our case report include considering HIT in CVST patients with an elevated 4T score and considering the entire clinical picture and degree of suspicion for HIT when interpreting negative HIT testing results. The PEA, in conjunction with the 4Ts score, may be considered as an alternate diagnostic assay for HIT.

Heparin-Induced Thrombocytopenia in Patients Undergoing Venoarterial Extracorporeal Membrane Oxygenation.

Background: Heparin-induced thrombocytopenia (HIT) is a serious, immune-mediated adverse drug reaction to unfractionated heparin (UFH) affecting also patients undergoing venoarterial extracorporeal membrane oxygenation (VA-ECMO). Although the association between VA-ECMO support and the development of thrombocytopenia has long been known and discussed, HIT as one underlying cause is still insufficiently understood. Therefore, the purpose of this study was to further investigate the epidemiology, mortality, diagnosis, and clinical management of HIT occurring in VA-ECMO patients treated with UFH. Methods: We conducted a retrospective single-center study including adult patients (≥18 years) with VA-ECMO support in the cardiac intensive care unit (ICU) of the University Hospital of Munich (LMU) between January 2013 and May 2022, excluding patients with a known history of HIT upon admission. Differences in baseline characteristics and clinical outcome between excluded HIT (positive anti-platelet factor 4 (PF4)/heparin antibody test but negative functional assay) and confirmed HIT (positive anti-PF4/heparin antibody test and positive functional assay) VA-ECMO patients as well as diagnosis and clinical management of HIT were analysed. Results: Among the 373 patients included, anti-PF4/heparin antibodies were detected in 53/373 (14.2%) patients. Functional HIT testing confirmed HIT in 13 cases (3.5%) and excluded HIT in 40 cases (10.7%), corresponding to a prevalence of confirmed HIT of 13/373 (3.5%) [1.6, 5.3] and a positive predictive value (PPV) of 24.5% for the antibody screening test. The platelet course including platelet recovery following argatroban initiation was similar between all groups. One-month mortality in patients with excluded HIT was 14/40 (35%) and 3-month mortality 17/40 (43%), compared to 5/13 (38%) (p > 0.999), and 6/13 (46%) (p > 0.999) in patients with confirmed HIT. Neurological outcome in both groups measured by the cerebral performance category of survivors on hospital discharge was similar, as well as adverse events during VA-ECMO therapy. Conclusions: With a prevalence of 3.5%, HIT is a non-frequent complication in patients on VA-ECMO and was not associated with a higher mortality rate. HIT was ultimately excluded by functional essay in 75% of VA-ECMO patients with clinical suspicion of HIT and positive anti-PF4/heparin antibody test. Argatroban seems to be an appropriate and safe therapeutic option for confirmed HIT-positive patients on VA-ECMO support.

Rethinking the Gold Standard: Real-World Performance of the Serotonin Release Assay (SRA) and PF4-Dependent P-Selectin Expression Assay (PEA) in the Diagnosis of Heparin-Induced Thrombocytopenia (HIT)

Background Heparin-induced thrombocytopenia (HIT) is a complication of heparin exposure with potential for significant morbidity and mortality. The serotonin release assay (SRA) has been considered the gold standard for HIT diagnosis. However, the SRA is technically challenging and results can be influenced by specimen handling and preparation. Therefore, the performance of the real-world assays may differ from published standards but reports of real-world performance are not widely available. A new assay, the PF4-dependent P-selectin expression assay (PEA), has demonstrated comparable accuracy in HIT diagnosis employing a simpler laboratory technique. Herein, we report the real-world performance of a reference laboratory SRA and PEA and describe how incorporation of the PEA into the testing algorithm positively influenced the HIT diagnostic process at a large academic medical center. Methods At the University of Alabama at Birmingham, we incorporated the PEA into HIT testing at provider discretion in patients with inconclusive SRA results starting January 1, 2020. Since July 15, 2020, we solidified the use of PEA in our algorithm (Figure 1) in which all patients with an inconclusive SRA had a PEA on the same specimen. A PEA could also be ordered ad hoc if the SRA result did not correlate with clinical suspicion for HIT. We retrospectively compared patients tested for HIT for 2 years before PEA use (January 1, 2018 to December 31, 2019; pre-PEA period) to the >2 years since using the PEA (January 1, 2020 to March 1, 2022; PEA period). All patients tested with PEA at our institution are included in Table 1. Laboratory testing: Enzyme immunoassay (EIA) was performed at our institution using Asserachrom® HPIA, Diagnostica Stago (France). The SRA and PEA assays were performed at Versiti (Milwaukee, WI). SRA was positive if there was >20% release of serotonin with low dose heparin (0.1 U/mL) and <20% release in with high dose heparin (100 U/mL). An inconclusive result was defined as a percentage of serotonin release not meeting criteria for a positive result (e.g., >20% release of serotonin with both low and high dose heparin, or some release but <20% with low dose along with inhibition with high dose). Statistical analysis Descriptive statistics were employed to examine the sample of patients with an inconclusive SRA result, comparing those with and without PEA testing. Bivariate analyses were conducted to compare testing patterns in the pre-PEA and PEA periods, and T-tests of significance were conducted to analyze assay results. All models were estimated using Stata 17 (StataCorp. 2019. Stata Statistical Software: Release 17. College Station, TX: StataCorp LLC). ResultsSRA results: In the pre-PEA and PEA periods, there were 1,252 and 1,403 EIAs performed, respectively. The rate of EIA positivity was similar in both groups (14% vs 15%, p=0.813). A total of 379 SRAs were sent in both periods, with 24 inconclusive results (6%, 15 pre-PEA and 9 PEA). PEA performance: In the 24 patients with inconclusive SRA results, HIT was diagnosed in 7 of 15 patients (47%) pre-PEA, compared to 3 of 9 patients (33%) in the PEA period. Table 1 presents the details of each case where PEA was used due to diagnostic uncertainty. Of the 9 patients with an inconclusive SRA result in the PEA period, PEA provided a definitive result in 8 cases (89%). In 2 cases, the PEA confirmed the SRA result, 1 positive and 1 negative. In 1 case with a positive SRA and low clinical suspicion, the PEA was negative. Conclusion Evaluation of the real-world performance of the SRA reveals that although inconclusive results are rare (6%), they can present a significant diagnostic challenge in centers with high testing volume. The PEA demonstrated reliable performance in patients with an inconclusive SRA, producing a definitive result in 89% of cases. The percentage of patients diagnosed with HIT after an inconclusive SRA result decreased with incorporation of the PEA (47% to 33%), suggesting that PEA use helped refute a HIT diagnosis when there was clinical uncertainty. Further studies are needed to determine the optimal testing algorithm to produce rapid, reliable HIT diagnosis. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

Genome-wide association study of platelet factor 4/heparin antibodies in heparin-induced thrombocytopenia

AbstractHeparin, a widely used anticoagulant, carries the risk of an antibody-mediated adverse drug reaction, heparin-induced thrombocytopenia (HIT). A subset of heparin-treated patients produces detectable levels of antibodies against complexes of heparin bound to circulating platelet factor 4 (PF4). Using a genome-wide association study (GWAS) approach, we aimed to identify genetic variants associated with anti-PF4/heparin antibodies that account for the variable antibody response seen in HIT. We performed a GWAS on anti-PF4/heparin antibody levels determined via polyclonal enzyme-linked immunosorbent assays. Our discovery cohort (n = 4237) and replication cohort (n = 807) constituted patients with European ancestry and clinical suspicion of HIT, with cases confirmed via functional assay. Genome-wide significance was considered at α = 5 × 10−8. No variants were significantly associated with anti-PF4/heparin antibody levels in the discovery cohort at a genome-wide significant level. Secondary GWAS analyses included the identification of variants with suggestive associations in the discovery cohort (α = 1 × 10−4). The top variant in both cohorts was rs1555175145 (discovery β = −0.112 [0.018], P = 2.50 × 10−5; replication β = −0.104 [0.051], P = .041). In gene set enrichment analysis, 3 gene sets reached false discovery rate-adjusted significance (q < 0.05) in both discovery and replication cohorts: “Leukocyte Transendothelial Migration,” “Innate Immune Response,” and “Lyase Activity.” Our results indicate that genomic variation is not significantly associated with anti-PF4/heparin antibody levels. Given our power to identify variants with moderate frequencies and effect sizes, this evidence suggests genetic variation is not a primary driver of variable antibody response in heparin-treated patients with European ancestry.

The Predictive Value of the 4Ts and HEP Score at Recommended Cutoffs in Patients With Mechanical Circulatory Support Devices

Despite the increasing utilization of mechanical circulatory support (MCS) devices, the 4Ts and heparin-induced thrombocytopenia (HIT) Expert Probability (HEP) scores have not been validated in patients with suspected HIT requiring MCS. A retrospective cohort study. At a tertiary university hospital. Adults with suspected HIT requiring any MCS. A diagnostic investigation of HIT. Of the 299 patients included, there were 374 diagnostic investigations of HIT, of which 32 (8.6%) were HIT-probable (heparin PF4 immunoassay optical density ≥1 or positive serotonin release assay). The 4Ts score ≥4 demonstrated a pretest sensitivity of 0.56 (95% confidence interval [CI]: 0.39-0.72) and specificity of 0.8 (95% CI: 0.75-0.83). The HEP score ≥3 demonstrated a pretest sensitivity of 0.31 (95% CI: 0.18-0.49) and specificity of 0.83 (95% CI: 0.79-0.87). The area under the receiver operating characteristic curve for the 4Ts and HEP scores were 0.68 (95% CI: 0.63-0.73) and 0.63 (95% CI: 0.59-0.68), respectively, and were not statistically different (p=0.21). In patients with an intra-aortic balloon pump, neither the 4Ts nor HEP score had discriminatory ability to differentiate probable HIT. The HEP score had no discriminatory ability in patients with concomitant MCS devices. The 4Ts and HEP scores have a modest predictive performance for probable HIT in patients requiring MCS devices. A low 4Ts or HEP score does not reliably rule out HIT in patients requiring MCS, and clinical suspicion for HIT should be investigated, utilizing laboratory tests in this population.

Performance evaluation of the STA ECA-II reagent for specific assay of anti-IIa activity of argatroban

Argatroban is indicated in the management of patients with suspicion of Heparin-Induced Thrombocytopenia. Biological monitoring of argatroban can be carried out using the activated partial thromboplastin time (APTT) or the measurement of the anti-IIa activity. However, APTT is susceptible to many interferences making its use unsuitable. The objective of this work is to define and verify the performance of a dosing protocol for the anti-IIa activity of the argatroban from the STA ECA-II reagent (Diagnostica Stago) and compare them to the Hemoclot Thrombin Inhibitor proposed by Hyphen Biomed based on diluted thrombin time. Calibration curve shows a good correlation between anti-IIa activity and the coloration level of the reaction medium over the concentration range studied (0 to 2μg/mL). The analytical performances obtained at the native dilution of 1/10th are excellent, both in terms of repeatability and reproducibility with a coefficient of variation below 10%. Automatic relaunch at 1/20th for sample concentrations greater than 2μg/mL does not impact this performance level. We obtain a limit of quantification below 0.2μg/mL. The comparison of the methods gave consistent and well-correlated results on a wide range of concentrations (0.3 to 4μg/mL). The use of the STA ECA-II reagent is a valuable alternative to APTT for biological monitoring of anticoagulant action of argatroban.

La thrombopénie induite par héparine : mise au point

Heparin-induced thrombocytopenia (HIT) is a serious complication of heparin therapy. It is due to the synthesis of antibodies most often directed against platelet factor 4 (FP4) modified by heparin (H). HIT is manifested by a platelet count fall, associated with a high risk of venous or arterial thrombosis. The diagnosis of HIT is based on the assessment of clinical probability (4Ts score or change in platelet count after cardiac surgery) and the demonstration of heparin-modified anti-FP4 antibodies (FP4/H). If the immunological tests are positive, functional tests should be performed. In case of suspicion of HIT, it is necessary to urgently stop heparin therapy, to perform a doppler ultrasound of the lower limbs, and to prescribe an alternative anticoagulation agent at a curative dose. Currently, danaparoid sodium and argatroban are authorized. The diagnosis and management of HIT remain complex and requires multidisciplinary collaboration.

Heparin‐induced thrombocytopenia: Construction of a pretest diagnostic score derived from the analysis of a prospective multinational database, with internal validation

BackgroundDiagnosis of heparin‐induced thrombocytopenia (HIT) requires pretest probability assessment and dedicated laboratory assays. ObjectiveTo develop a pretest score for HIT. DesignObservational; analysis of prospectively collected data of hospitalized patients suspected with HIT (ClinicalTrials.gov NCT00748839). SettingThirty‐one tertiary hospitals in France, Switzerland, and Belgium. PatientsPatients tested for HIT antibodies (2280 evaluable), randomly allocated to derivation and validation cohorts. MeasurementsIndependent adjudicators diagnosed HIT based on the prospectively collected data and serotonin release assay results. ResultsHeparin‐induced thrombocytopenia was diagnosed in 234 (14.7%) and 99 (14.5%) patients in the two cohorts. Eight features were associated with HIT (in brackets, points assigned for score calculation of the score): unfractionated heparin (1); therapeutic‐dose heparin (1); cardiopulmonary bypass (cardiac surgery) (2); major trauma (3); 5‐ to 21‐day interval from anticoagulation initiation to suspicion of HIT (4); ≥40% decrease in platelet count over ≤6 days (3); thrombotic event, arterial (3) or venous (3). The C‐statistic was 0.79 (95% CI, 0.76–0.82). In the validation cohort, the area under the receiver operating characteristic curve was 0.77 (95% CI, 0.74–0.80). Three groups of scores were defined; HIT prevalence reached almost 30% in the high‐probability group. LimitationThe performance of the score may depend on settings and practices. ConclusionThe objective, easy‐to‐collect, clinical features of HIT we evidenced were incorporated into a pretest score, which may guide clinical decisions regarding diagnostic testing and anticoagulation.

Incidence of Heparin Induced Thrombocytopenia in Patients Undergoing Implantation of Durable and Non-Durable Mechanical Circulatory Support Devices

Purpose Unfractionated heparin exposure in patients undergoing mechanical circulatory support device (MCSD) implantation increases the risk for developing heparin-induced-thrombocytopenia (HIT). The false positive rate of HIT in this population using current standard testing is unknown. Postoperative and device related thrombocytopenia complicate the HIT diagnosis. Methods A retrospective analysis of patients undergoing MCSD pre-implantation assessment or implantation of MCSD (October 2014 - August 2020) was performed. The primary objective was to assess the incidence of HIT with positive PF4 IgG ELISA (PF4) and serotonin release (SRA) assays. False positive HIT was defined as PF4 ≥0.4 optical density (OD) and negative SRA. Argatroban usage was also assessed. Results 366 patients with 512 tests were analyzed. PF4 tests were performed in 128 patients (158 tests) undergoing pre-implantation assessment, 136 patients (185 tests) with non-durable devices (primarily Impella®), and 102 (169 tests) patients with durable devices (72 ventricular assist devices, 30 total artificial hearts). 25% (92/366) of patients had positive PF4 ≥0.4 OD and had subsequent SRA performed (92 patients, 122 tests). True positive HIT was documented in 13% (12/92) of PF4 positive patients consisting of 1 pre-implantation, 9 non-durable MCSD, and 2 durable MCSD patients. The mean PF4 for true positive HIT was 2.2±0.7 OD. False positive HIT was documented in 15% (19/128) of pre-implantation patients (mean PF4 0.9±0.6 OD), 14% (19/136) of non-durable MCSD patients (mean PF4 0.8±0.5 OD), and 41% (42/102) of durable MCSD patients (mean PF4 1.1±0.6 OD). Argatroban was administered in 40% (37/92) of patients with a positive PF4 and 67% (25/37) of patients with a negative SRA. Conclusion Without clinical suspicion of HIT, preoperative PF4 testing is not recommended. Patients with non-durable MCSD devices are at the highest risk of developing HIT (7%). Although a PF4 ≥0.4 OD is considered a positive screening test for HIT, a higher threshold should be considered for MCSD patients. To our knowledge, this is the first in depth assessment of the incidence of HIT in patients undergoing pre-implantation evaluation for both non-durable and durable MCS devices.

Clinical validation of immunoassay HemosIL® AcuStar HIT-IgG (PF4-H) in the diagnosis of Heparin-induced thrombocytopenia

Heparin induced thrombocytopenia (HIT) is a life and limb-threatening complication of heparin exposure. The misdiagnosis of this disease can have major consequences on the patients. The objective of this study was to evaluate a diagnostic strategy that combines the 4Ts score with the result of HemosIL® AcuStar HIT-IgG (PF4-H) to confirm the diagnosis of HIT. Citrated plasmas from 1300 patients with suspicion of HIT were analyzed with a fully automated quantitative chemiluminescent immunoassay (HemosIL® AcuStar HIT-IgG (PF4/H)). If the IgG anti-PF4/H antibodies were positive (cut-off, 1 U/mL), HIT diagnosis was confirmed using functional tests. In total, 1300 samples of consecutive patients were enrolled, 94 (7.2%) of which gave positive results in HemosIL® AcuStar-IgG. HIT was diagnosed in 65 out of these patients, corresponding to a prevalence of 5%. Using ROC curve analysis, patients were divided into three groups according to their titer of antibodies. Higher values of the IgG (PF4-H) were associated with increased probability of HIT, and the diagnostic specificity was greatly increased using the combination of a 4Ts score > 3 and a positive titer ≥ 3.25 U/mL. Importantly, the diagnostic specificity is 100% when the titer is > 12.40 U/mL. We demonstrated that higher values of Anti PF4/H Antibodies were associated with a high probability of having HIT. A titer of HemosIL® IgG (PF4-H) > 12.40 U/mL has a specificity of 100% which should no require a functional test to confirm the diagnosis of HIT.

Bleeding and Thrombotic Adverse Events in Hospitalized Patients Under Empiric Treatment for Suspected Heparin-Induced Thrombocytopenia While Awaiting Confirmatory Testing.

Empiric management in suspected heparin-induced thrombocytopenia (HIT) is challenging due to imperfect prediction models, latency while awaiting test results and risks of empiric therapies. When there is high clinical suspicion for HIT, cessation of heparin and empiric non-heparin anticoagulation with FDA-approved argatroban is recommended. Alternatively off-label fondaparinux or watchful waiting have been utilized in clinical practice. Outcomes of patients empirically managed for HIT have not been compared directly in clinical trials and patients that ultimately do not have HIT are often overlooked. Clinicians need studies investigating empiric management to guide decision making in suspected HIT. In this study, adverse events (AE) were categorized and compared in patients being evaluated for HIT while undergoing empiric management by non-heparin anticoagulation with argatroban or fondaparinux, both at therapeutic or reduced doses, or watchful waiting with or without heparin. AE were defined as new thrombosis confirmed on imaging or new bleeding event after HIT was first suspected. A retrospective chart review of 312 patients tested for HIT at an academic hospital was conducted. 170 patients met inclusion criteria. Patients were excluded if the 4Ts score was < 4. The 4Ts score is a pretest probability for HIT based on thrombocytopenia degree, timing, alternative causes and presence of thrombosis. Included patients were divided according to management groups and compared with logistic regression analysis. Bleeding risk significantly differed between management groups (p = 0.002). Despite adjustment for bleeding risk, fondaparinux was associated with increased AE, (p = 0.03, OR = 5.81), while argatroban was not. There was no difference in AE based on time to initiation of empiric treatment and no advantage to reduced dosing with either anticoagulant. These findings challenge assumptions surrounding empiric HIT management.

Heparin Induced Thrombocytopenia in Patients with COVID-19

BackgroundThe Coronavirus disease-2019 (COVID-19) is a global pandemic caused by novel coronavirus SARS-CoV-2. Acute respiratory and renal failure as well as systemic coagulopathy are critical aspects of the morbidity and mortality in patients with COVID-19. Heparin Induced Thrombocytopenia (HIT) occurs when IgG antibodies form against platelet factor 4-Heparin complex, resulting in platelet activation and removal, leading to a prothrombotic state. HIT is suspected when there is a platelet count decrease of more than 50% after exposure to heparin, along with hypercoagulability. Clinical Scoring systems like 4T Score (Thrombocytopenia, Timing, Thrombosis, no other cause of Thrombocytopenia) have been developed to assess the pretest probability of HIT. The use of unfractionated heparin, post-orthopedic and post-cardiac surgery state, female gender, and old age are recognized as risk factors for HIT. There is a nine-fold increased risk of developing HIT in patients requiring hemodialysis. ICU patients and patients with VTE without thrombocytopenia are considered to have low pre-test probability for HIT. Studies have shown that only 6% who are investigated serologically for HIT actually have the diagnosis. We conducted this study to assess the incidence and risk factors for HIT in COVID-19 positive patients and its impact on mortality.MethodsA retrospective analysis was performed on all patients who were COVID-19 positive and hospitalized between March 1, 2020 and June 26, 2020 at our institution. Patients with intermediate or high suspicion for HIT, based on 4T score of 4 or higher, underwent IgG-specific platelet factor 4(PF4)-dependent enzyme immune assay (EIA). Washed platelet assays such as serotonin release assay (SRA) and heparin-induced platelet aggregation (HIPA) were used as confirmatory tests in cases with intermediate or low optical density (OD) with EIA. The incidence of HIT, its impact on mortality, and positivity of IgG-specific PF4-EIA in COVID-19 patients were studied, and statistical analysis was done with X2 testing. Subgroup analysis was performed based on demographic factors and risk factors for HIT, including exposure to heparin or low molecular weight heparin (LMWH), history of or cancer, recent orthopedic or cardiac surgery, exposure to renal replacement therapy (RRT), and severity of disease (D-dimer >6 ULN, Acute Kidney Injury, ICU admission, and mechanical ventilation requirement). These factors were analyzed by Fisher’s exact test to determine their impact on mortality. The hospital course for HIT antibody-positive patients was further analyzed to study the impact of COVID-19 related therapy, such as Remdesivir, Tocilizumab, Hydroxychloroquine, Steroids, and anticoagulation after diagnosis of HIT.ResultsWEight out of 1265 hospitalized COVID-19 positive patients tested positive for IgG-specific platelet factor 4(PF4)-dependent enzyme immune assay (EIA+). Incidence of EIA+ in COVID-19 patients was 0.6%, which is significantly higher than in the general population 0.2% (p<.0001, 95% CI 0.25-1.20%). The incidence of thromboembolic events in EIA+ patients was 87.5%, significantly higher than the rate of 10.90% in all COVID-19 patients (p<.0001, CI 41.96- 86.98%). The mortality rate in EIA+ patients was 50%, significantly greater than the mortality rate of 12% in all hospitalized COVID-19 patients (p=.0011, CI 9.46-66.53). Serological confirmation of HIT diagnosis was 37.5% (2 had OD>1, 1 was SRA positive) which is significantly higher than confirmation of HIT in nonCOVID-19 patients 6% (p<.0001, 95% CI 29.57-85.32%). All 3 confirmed HIT patients (100%) had severe disease (3/3 required ICU admission, 2/3 required mechanical ventilation, the one not requiring mechanical ventilation required RRT).ConclusionOur study indicates incidence of HIT is higher in the COVID-19 population. The incidence of positive EIA for patients with intermediate to high 4T scores is also higher in COVID-19 positive patients. This can be attributed to the cytokine storm and severe sepsis seen in critically ill COVID-19 patients. Our study also suggests that development of HIT can contribute to increased risk for thromboembolic events as well as mortality of COVID-19 patients, however, our study is limited due to small sample size. Therefore, prospective studies are needed to analyze the impact of HIT on morbidity, mortality and long-term outcomes in COVID-19 patients.TableDisclosuresNo relevant conflicts of interest to declare.